Nephronophthisis (NPHP) is an important public health issue because it is the most common genetic cause of end-stage renal disease in the first three decades of life. The kidneys of patients with NPHP demonstrate scarring and cyst formation, features shared with many other types of kidney diseases. The study of nephrocystin-5 will provide insight into the mechanisms for kidney scarring and cyst formation in NPHP. These mechanisms may be common to other causes of kidney disease and kidney failure. The sponsor's lab has identified by positional cloning mutations in three novel genes (NPHP1, 2, and 4) to cause NPHP. They also recently identified mutations in the inversin gene as causing NPHP type 2, thus linking renal cystic disease to the function of primary cilia and left-right axis determination (Otto et al. Nature Genet 34:413, 2003). Very recently another novel gene, NPHP5, was identified in patients with NPHP type 5 and retinitis pigmentosa (Otto, et al. Nature Genet. 37:282-8, 2005). The gene product of NPHP5 is nephrocystin-5. The overall hypothesis of this application is that nephrocystin-5 participates in a protein complex, which likely includes the other nephrocystin proteins, and functions in the formation of tight cell junctions and apical-basal polarization. Specifically we will: 1. Examine the subcellular localization of the novel protein, nephrocystin-5. We will employ laser scanning confocal microscopy to examine cell in tissue culture as well as murine tissue sections. 2. Examine how nephrocystin-5 participates in a functional protein complex. We will examine the nephrocystin-5 protein complex with co-immunoprecipitations, 2D-gels and mass spectrometry. 3. Characterize the effect of NPHP5 mutations on the subcellular localization of nephrocystin-5, the integrity of the epithelial monolayer and cell polarization. MDCK cells expressing truncated forms of nephrocystin-5 will be generated. These cells will be evaluated for the localization of nephrocystin-5 with confocal microscopy, assayed for the formation of tight cell junctions with transepithelial electrical resistance and evaluated for polarization defects by growing them in a collagen gel.